Eicosanoid-Regulated Myeloid ENaC and Isolevuglandin Formation in Human Salt-Sensitive Hypertension.

Background: The mechanisms by which salt increases blood pressure in people with salt sensitivity remain unclear. Our previous studies found that high sodium enters antigen-presenting cells (APCs) via the epithelial sodium channel and leads to the production of isolevuglandins and hypertension. In the current mechanistic clinical study, we hypothesized that epithelial sodium channel-dependent isolevuglandin-adduct formation in APCs is regulated by epoxyeicosatrienoic acids (EETs) and leads to salt-sensitive hypertension in humans.

Impaired Dynamic Sarcoplasmic Reticulum Ca Buffering in Autosomal Dominant CPVT2.

Background: Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a potentially lethal cardiac arrhythmia syndrome triggered by catecholamines released during exercise, stress, or sudden emotion. Variants in the calsequestrin-2 gene (), encoding the major calcium (Ca) binding protein in the sarcoplasmic reticulum (SR), are the second most common cause of CPVT. Recently, several gene variants, such as -K180R, have been linked to an autosomal dominant form of Casq2-linked CPVT (CPVT2), but the underlying mechanism is not known.

RYR2 Channel Inhibition Is the Principal Mechanism of Flecainide Action in CPVT.

Rationale: The class Ic antiarrhythmic drug flecainide prevents ventricular tachyarrhythmia in patients with catecholaminergic polymorphic ventricular tachycardia (CPVT), a disease caused by hyperactive RyR2 (cardiac ryanodine receptor) mediated calcium (Ca) release. Although flecainide inhibits single RyR2 channels in vitro, reports have claimed that RyR2 inhibition by flecainide is not relevant for its mechanism of antiarrhythmic action and concluded that sodium channel block alone is responsible for flecainide's efficacy in CPVT.

Objective: To determine whether RyR2 block independently contributes to flecainide's efficacy for suppressing spontaneous sarcoplasmic reticulum Ca release and for preventing ventricular tachycardia in vivo.

Patient-independent human induced pluripotent stem cell model: A new tool for rapid determination of genetic variant pathogenicity in long QT syndrome.

Background: Commercial genetic testing for long QT syndrome (LQTS) has rapidly expanded, but the inability to accurately predict whether a rare variant is pathogenic has limited its clinical benefit. Novel missense variants are routinely reported as variant of unknown significance (VUS) and cannot be used to screen family members at risk for sudden cardiac death. Better approaches to determine the pathogenicity of VUS are needed.

Unnatural verticilide enantiomer inhibits type 2 ryanodine receptor-mediated calcium leak and is antiarrhythmic.

Ca leak via ryanodine receptor type 2 (RyR2) can cause potentially fatal arrhythmias in a variety of heart diseases and has also been implicated in neurodegenerative and seizure disorders, making RyR2 an attractive therapeutic target for drug development. Here we synthesized and investigated the fungal natural product and known insect RyR antagonist (-)-verticilide and several congeners to determine their activity against mammalian RyR2. Although the cyclooligomeric depsipeptide natural product (-)-verticilide had no effect, its nonnatural enantiomer [-(+)-verticilide] significantly reduced RyR2-mediated spontaneous Ca leak both in cardiomyocytes from wild-type mouse and from a gene-targeted mouse model of Ca leak-induced arrhythmias (2).

Hypertrophic cardiomyopathy-linked mutation in troponin T causes myofibrillar disarray and pro-arrhythmic action potential changes in human iPSC cardiomyocytes.

Background: Mutations in cardiac troponin T (TnT) are linked to increased risk of ventricular arrhythmia and sudden death despite causing little to no cardiac hypertrophy. Studies in mice suggest that the hypertrophic cardiomyopathy (HCM)-associated TnT-I79N mutation increases myofilament Ca sensitivity and is arrhythmogenic, but whether findings from mice translate to human cardiomyocyte electrophysiology is not known.

Objectives: To study the effects of the TnT-I79N mutation in human cardiomyocytes.

Dendritic Cell Amiloride-Sensitive Channels Mediate Sodium-Induced Inflammation and Hypertension.

Sodium accumulates in the interstitium and promotes inflammation through poorly defined mechanisms. We describe a pathway by which sodium enters dendritic cells (DCs) through amiloride-sensitive channels including the alpha and gamma subunits of the epithelial sodium channel and the sodium hydrogen exchanger 1. This leads to calcium influx via the sodium calcium exchanger, activation of protein kinase C (PKC), phosphorylation of p47, and association of p47 with gp91.

Thyroid and Glucocorticoid Hormones Promote Functional T-Tubule Development in Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

Rationale: Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) are increasingly being used for modeling heart disease and are under development for regeneration of the injured heart. However, incomplete structural and functional maturation of hiPSC-CM, including lack of T-tubules, immature excitation-contraction coupling, and inefficient Ca-induced Ca release remain major limitations.

Objective: Thyroid and glucocorticoid hormones are critical for heart maturation.

Novel CPVT-Associated Calmodulin Mutation in CALM3 (CALM3-A103V) Activates Arrhythmogenic Ca Waves and Sparks.

Background: Calmodulin (CaM) mutations are associated with severe forms of long QT syndrome and catecholaminergic polymorphic ventricular tachycardia (CPVT). CaM mutations are found in 13% of genotype-negative long QT syndrome patients, but the prevalence of CaM mutations in genotype-negative CPVT patients is unknown. Here, we identify and characterize CaM mutations in 12 patients with genotype-negative but clinically diagnosed CPVT.

Spectrum and Prevalence of CALM1-, CALM2-, and CALM3-Encoded Calmodulin Variants in Long QT Syndrome and Functional Characterization of a Novel Long QT Syndrome-Associated Calmodulin Missense Variant, E141G.

Background: Calmodulin (CaM) is encoded by 3 genes, CALM1, CALM2, and CALM3, all of which harbor pathogenic variants linked to long QT syndrome (LQTS) with early and severe expressivity. These LQTS-causative variants reduce CaM affinity to Ca(2+) and alter the properties of the cardiac L-type calcium channel (CaV1.2).

In Vitro Studies Indicate Intravenous Lipid Emulsion Acts as Lipid Sink in Verapamil Poisoning.

Intravenous lipid emulsion (ILE), a component of parenteral nutrition, consists of a fat emulsion of soy bean oil, egg phospholipids, and glycerin. Case reports suggest that ILE may reverse hypotension caused by acute poisoning with lipophilic drugs such as verapamil, but the mechanism remains unclear. The methods used are the following: (1) measurement of ILE concentration in serum samples from a patient with verapamil poisoning treated with ILE, (2) measurement of free verapamil concentrations in human serum mixed in vitro with increasing concentrations of ILE, and (3) measurement of murine ventricular cardiomyocyte L-type Ca(2+) currents, intracellular Ca(2+), and contractility in response to verapamil and/or ILE.

Comparable calcium handling of human iPSC-derived cardiomyocytes generated by multiple laboratories.

Cardiomyocytes (CMs) derived from human induced pluripotent stem cells (hiPSCs) are being increasingly used to model human heart diseases. hiPSC-CMs generated by earlier aggregation-based methods (i.e.

Impaired calcium-calmodulin-dependent inactivation of Cav1.2 contributes to loss of sarcoplasmic reticulum calcium release refractoriness in mice lacking calsequestrin 2.

Aims: In cardiac muscle, Ca(2+) release from sarcoplasmic reticulum (SR) is reduced with successively shorter coupling intervals of premature stimuli, a phenomenon known as SR Ca(2+) release refractoriness. We recently reported that the SR luminal Ca(2+) binding protein calsequestrin 2 (Casq2) contributes to release refractoriness in intact mouse hearts, but the underlying mechanisms remain unclear. Here, we further investigate the mechanisms responsible for physiological release refractoriness.

Coordinated regulation of murine cardiomyocyte contractility by nanomolar (-)-epigallocatechin-3-gallate, the major green tea catechin.

Green tea polyphenolic catechins exhibit biological activity in a wide variety of cell types. Although reports in the lay and scientific literature suggest therapeutic potential for improving cardiovascular health, the underlying molecular mechanisms of action remain unclear. Previous studies have implicated a wide range of molecular targets in cardiac muscle for the major green tea catechin, (-)-epigallocatechin-3-gallate (EGCG), but effects were observed only at micromolar concentrations of unclear clinical relevance.

Myofilament Ca sensitization increases cytosolic Ca binding affinity, alters intracellular Ca homeostasis, and causes pause-dependent Ca-triggered arrhythmia.

Rationale: Ca binding to the troponin complex represents a major portion of cytosolic Ca buffering. Troponin mutations that increase myofilament Ca sensitivity are associated with familial hypertrophic cardiomyopathy and confer a high risk for sudden death. In mice, Ca sensitization causes ventricular arrhythmias, but the underlying mechanisms remain unclear.

Calsequestrin mutations and catecholaminergic polymorphic ventricular tachycardia.

Cardiac calsequestrin (Casq2) is the major Ca2+ binding protein in the sarcoplasmic reticulum, which is the principle Ca2+ storage organelle of cardiac muscle. During the last decade, experimental studies have provided new concepts on the role of Casq2 in the regulation of cardiac muscle Ca2+ handling. Furthermore, mutations in the gene encoding for cardiac calsequestrin, CASQ2, cause a rare but severe form of catecholaminergic polymorphic ventricular tachycardia (CPVT).

Role of intracellular stores in the regulation of rhythmical [Ca2+]i changes in interstitial cells of Cajal from rabbit portal vein.

Interstitial cells of Cajal (ICCs) freshly isolated from rabbit portal vein and loaded with the Ca(2+)-sensitive indicator fluo-3 revealed rhythmical [Ca(2+)](i) changes occurring at 0.02-0.1 Hz.

Voltage-gated potassium currents in rat vas deferens smooth muscle cells.

Voltage-gated components of the outward current in single smooth muscle cells isolated from the epididymal part of the rat vas deferens were studied using amphotericin B perforated patch-clamp techniques. The complex kinetics of the net outward current elicited by positive voltage steps from -80 mV to +40 mV suggested the presence of several components. Bath application of 200 nM charybdotoxin, a potent blocker of large-conductance, Ca(2+)-dependent K(+) channels (BK(Ca)), reduced the current amplitude significantly.